Ghrelin production promoter

ABSTRACT

The objectives are to find a substance or a composition capable of promoting the production of ghrelin with physiological activities such as growth hormone secretion effect, and to provide a pharmaceutical comprising it as the active ingredient. The pharmaceutical is a ghrelin production promoter comprising Rikkunshi-to as the active ingredient.

TECHNICAL FIELD

The present invention relates to a novel pharmaceutical such as ghrelinproduction promoter comprising “Rikkunshi-to” (a kind of kampo medicine)as the active ingredient.

BACKGROUND ART

Ghrelin is a peptide hormone discovered in 1999 that is known to promotethe secretion of growth hormone (GH). Ghrelin promotes GH secretion viareceptors different from those for growth hormone-releasing hormone(GHRH). The peptide chain of ghrelin comprises 28 amino acid residues,in which the hydroxyl group of the third serine is acylated withoctanoic acid (active ghrelin); but non-acylated ghrelin (desacylghrelin) does not exhibit GH secretion activity.

Various physiological active mechanisms of ghrelin are known; and forexample, ghrelin is known to increase appetite, protect thecardiovascular system, and control metabolism among other effects, inaddition to the above-mentioned GH secretion-promoting effect. Theseknown effects of ghrelin suggest that the possibility of its applicationto therapeutic agents for diseases such as cibophobia, myocardialinfarction, heart failure, and cachexia as well as those to preventaging/dwarfism (Non-Patent Document 1). Moreover, it is reported thatghrelin promotes neurite extension and improves nerve paralysis(including movement disorders and perceptual disorders) associated withperipheral nerve diseases such as amyotrophic lateral sclerosis,diabetic neurological disorder, neuropathy, neurological disorder causedby traumatic neural damage or neural deficiency, toxic neurologicaldisorder and multiple sclerosis. It also improves memory disorders inpatients with central nerve disorders and diseases such as Alzheimer'sdisease, vascular dementia, Parkinson's disease, Huntington's diseaseand spinal injury; it is useful in the treatment of dementia and forbrain function activation; as well as to treat and prevent amyotrophiclateral sclerosis and diabetic neurological disorder (Patent Document1). It is also known for its activation of osteoblasts and bonereconstitution in dwarfism and normal persons; increase of muscle massand muscle strength in GH-deficient adults; improvement of exercisecapacity in GH-deficient adults; and cure of severe burns in children.It is used in combination with gonadotropin to induce ovulation; in theprevention of protein metabolism disorder due to prednisoneadministration; in the promotion of T-cell education in severeimmunodeficiency; and prevention of senile weight loss, fatty tissueextension and skin atrophy. As for the effects of ghrelin in patientswith disorders not directly related to GH deficiency or depression,ghrelin may, for example, increase heart beating and is therefore knownto be effective in cardiac diseases such as heart failure (PatentDocument 2). Also known are its effects on myocardial infarction,reperfusion injury in bypass surgery or PTCA for myocardial infarction,coronary microcirculation failure, myocarditis (alcoholic, viral, etc.),dilated cardiomyopathy, cardiac transplantation, arrhythmia, heartfailure, and also drug-induced myocardial disorders caused bychemotherapeutical agents or the like (Patent Document 3). In addition,it has been confirmed that ghrelin administration to patients withchronic obstructive pulmonary disease (COPD) results in an increase ofrespiration muscle power, body weight and skeletal muscle mass, as wellas in an improvement of the QOL score and the 6-minute walking distance;it is also reported that ghrelin may be expected to improve varioussymptoms of COPD (Non-Patent Document 2).

As in the above, various excellent physiological effects of ghrelin havebeen specifically noted, and the development of pharmaceuticalscomprising ghrelin as the active ingredient has been promoted. However,few substances have heretofore been known capable of promoting ghrelinproduction in living bodies.

Patent Document 1: JP-A 2005-239712

Patent Document 2: Re-issued WO03097083

Patent Document 3: Re-issued WO04014412

Non-Patent Document 1: Proceedings the 124th Symposium of the JapaneseAssociation of Medical Science, pp. 45-52

Non-Patent Document 2: New Current 16 (27), pp. 17-22

DISCLOSURE OF THE INVENTION Problems to be Solved by the Invention

The present invention has been made in consideration of the technicalbackground described above, and its objectives are to find a substanceor composition that will promote ghrelin production in living bodies andto provide a pharmaceutical comprising it as the active ingredient.

Means for Solving the Problems

The present inventors have assiduously investigated substances andcompositions that may promote ghrelin production. In the end, they foundthat “Rikkunshi-to”, a kind of kampo medicine has an excellent ghrelinproduction-promoting effect, and have completed the present invention.

Specifically, the invention is a ghrelin production promoter comprisingRikkunshi-to as the active ingredient.

EFFECT OF THE INVENTION

According to the invention, it is possible to provide a pharmaceutical,which may effectively increase the production of acylated active ghrelinand be an effective treatment for patients with disorders associatedwith growth hormone deficiency or depression such as dwarfism, centralnerve disorders such as Alzheimer's disease, and cardiac disorders suchas myocardial infarction.

The pharmaceutical of the invention has an excellent safety profile withfew problems such as side effects, since the active ingredientRikkunshi-to has been used traditionally as a prescription drug in Japanand China.

BEST MODE FOR CARRYING OUT THE INVENTION

Rikkunshi-to used as the active ingredient in the invention is a type ofkampo medicine that contains crude drugs such as ATRACTYLODIS LANCEAERHIZOMA, GINSENG RADIX, PINELLIAE TUBER, POLIA, ZIZYPHI FRUCTUS andothers described in the classics of kampo medicine formulation (ManbyoKaishun); it promotes gastric emptying and increase gastric mucosalblood flow, and is used as a therapeutic agent for gastritis and gastricatony.

The compounding range (composition) of the crude drugs in theRikkunshi-to is generally as follows:

ATRACTYLODIS LANCEAE RHIZOMA (JP Atractylodes Lancea Rhizome) orATRACTYLODIS RHIZOMA (JP Atractylodes Rhizome) 3.0 to 4.0 GINSENG RADIX(JP Ginseng) 2.0 to 4.0 PINELLIAE TUBER (JP Pinellia Tuber) 3.0 to 4.0POLIA (JP Poria Sclerotium) 3.0 to 4.0 ZIZYPHI FRUCTUS (JP Jujube) 2.0AURANTII NOBILIS PERICARPIUM (JP Citrus Unshiu Peel) 2.0 to 4.0GLYCYRRHIZAE RADIX (JP Glycyrrhiza) 1.0 to 1.5 ZINGIBERIS RHIZOMA (JPGinger) 0.5 to 2.0

Of the compounding range, the especially preferred Rikkunshi-to isobtained from a mixture of the crude drugs of ATRACTYLODIS LANCEAERHIZOMA 4.0, GINSENG RADIX 4.0, PINELLIAE TUBER 4.0, POLIA 4.0, ZIZYPHIFRUCTUS 2.0, AURANTII NOBILIS PERICARPIUM 2.0, GLYCYRRHIZAE RADIX 1.0and ZINGIBERIS RHIZOMA 0.5.

In the ghrelin production promoter of the invention, either Rikkunshi-tohaving the above-mentioned composition itself or its extract can be usedas the active ingredient of a pharmaceutical formulation prepared usingknown carriers for pharmaceuticals.

The extract of Rikkunshi-to can be prepared according to classicalmethods in which a mixture of the crude drugs mixed at the compoundingratio mentioned above is extracted with 5 to 20 volumes of anappropriate aqueous solvent followed by solid-liquid separation.Examples of the preferred aqueous solvent are water, ethanol and aqueoussolution of acetic acid. The extraction method can be hot or coldextraction. In particular, extraction conducted at 90 to 100° C. usingwater as the solvent is preferred.

The obtained extract of Rikkunshi-to may be dried into powder ifnecessary. Although the extract, in liquid or powder state, can bedirectly administered as it is, it may also be formulated into anextract preparation for easy administration and convenient carrying. Forexample, a mixture of the crude drugs is extracted with 10 volumes ofhot water followed by solid-liquid separation. The obtained liquidextract is concentrated and dried, and the resulting dried powderextract of Rikkunshi-to is formulated as diluted powder, granules,tablets or capsules according to conventional methods by mixing it withappropriate fillers (such as lactose, corn starch and crystallinecellulose), excipients or the like, which are commonly used forpharmaceutical formulations.

The ghrelin production promoter of the invention may be administered asany formulation for oral use, or for parenteral use such as injection orinfusion.

Formulations for oral use include solid ones such as powder, granules,tablets and capsules, and liquid ones such as suspensions, emulsions,syrups, and elixirs; and pharmaceutical carriers may be used inaccordance with the administration routes and formulations.

As for solid formulations for oral use, the usable carriers are starch,lactose, sugar, mannitol, carboxymethyl cellulose, corn starch,inorganic salts, etc. Furthermore, optional components such as binders,disintegrating agents, surfactants, lubricants, fluidity accelerators,corrigents, coloring agents and perfumes may be compounded as needed.

Of the above optional components, examples of the binders includestarch, dextrin, acacia, gelatin, hydroxypropyl starch, methylcellulose, sodium carboxymethyl cellulose, hydroxypropyl cellulose,crystalline cellulose, ethyl cellulose, polyvinyl pyrrolidone, andmacrogol.

Examples of the disintegrating agents include starch,hydroxypropylstarch, sodium carboxymethyl cellulose, calciumcarboxymethyl cellulose, carboxymethyl cellulose and low-substitutedhydroxypropyl cellulose.

Examples of the surfactants include sodium laurylsulfate, soybeanlecithin, sucrose fatty acid ester and Polysorbate 80. Examples of thelubricants include talc, waxes, hydrogenated vegetable oil, sucrosefatty acid ester, magnesium stearate, calcium stearate, aluminiumstearate, polyethylene glycol. Examples of the fluidity acceleratorsinclude light silicic anhydride, dried aluminium hydroxide gel,synthetic aluminium silicate and magnesium silicate.

As for formulations for parenteral use, these may be produced asfollows: The active ingredient, Rikkunshi-to or its extract is dissolvedor suspended in distilled water for injection, physiological salinewater, aqueous sucrose water, vegetable oil for injection, sesame oil,peanut oil, soybean oil, corn oil, propylene glycol, polyethylene glycolor the like according to conventional methods; if desired, amicrobicide, a preservative, a stabilizer, a tonicity agent, ananalgesic or the like, may be added thereto.

The dose of the ghrelin production promoter of the invention describedabove may vary depending on the administrating route, the diseasecondition and others; but in general, for oral administration, the dosemay be from 500 to 1000 mg/kg/day or so of Rikkunshi-to extract.

As already described, ghrelin is effective for activation of osteoblastsand bone reconstitution inpatients with dwarfism and in normal persons,increases muscle mass and muscle strength in GH-deficient adults, andimproves in exercise capacity of GH-deficient adults, etc. Therefore,the ghrelin production promoter comprising Rikkunshi-to as the activeingredient may be used as a growth hormone secretion promoter.

Ghrelin is known to contribute to treatment of myocardial infarction,reperfusion injury after bypass surgery or PTCA for myocardialinfarction, coronary microcirculation failure, myocarditis (alcoholic,viral, etc.), dilated cardiomyopathy, cardiac transplantation,arrhythmia, heart failure, and also myocardial disorders induced bydrugs such as chemotherapeutical agents or the like. Consequently, theghrelin production promoter comprising Rikkunshi-to as the activeingredient may be used as a therapeutic agent inpatients with myocardialinfarction or heart failure.

Moreover, it is reported that ghrelin promotes neurite extension andimproves nerve paralysis (including movement disorders and perceptualdisorders) associated with peripheral nerve diseases such as amyotrophiclateral sclerosis, diabetic neurological disorder, neuropathy,neurological disorder caused by traumatic neural damage or neuraldeficiency, toxic neurological disorder and multiple sclerosis. It alsoimproves memory disorders in patients with central nerve disorders anddiseases such as Alzheimer's disease, vascular dementia, Parkinson'sdisease, Huntington's disease and spinal injury; it is useful in thetreatment of dementia and for brain function activation; as well as totreat and prevent amyotrophic lateral sclerosis; the ghrelin productionpromoter comprising Rikkunshi-to as the active ingredient may be used asa therapeutic agent for patients with Alzheimer's disease.

Besides, it is reported that ghrelin administration to patients withchronic obstructive pulmonary disease (COPD) results in an increase ofrespiration muscle power, body weight and skeletal muscle mass, as wellas in an improvement of the QOL score and the 6-minute walking distance;and therefore, the ghrelin production promoter comprising Rikkunshi-toas the active ingredient may be used as a therapeutic agent for patientswith chronic obstructive pulmonary disease.

EXAMPLES

The invention is described in more detail with reference to thefollowing Examples, however, the invention is not restricted at all bythese Examples.

Test Example 1 Ghrelin Production Promotion Test 1

The ghrelin production promoting effect was investigated in fastednormal rats without anticancer agent treatment. Seven-week-old SD malerats (8/group) were orally administered a test substance Rikkunshi-to(TJ-43, by Tsumura & Co.) at a dose of 100 or 500 mg/kg, and kept fastedfor 24 hours. Then, Rikkunshi-to was orally administered again at thesame dose and two hours later the rats were decapitated and the bloodwas collected to determine the concentration of ghrelin by ELISA. Bloodwas centrifuged at 10,000 rpm for three minutes, then 1 mol HCl wasadded to the collected supernatant in proportion of 1/10 of the blood toprepare the test sample. The ghrelin concentration was quantitativelydetermined using an Active Ghrelin ELISA Kit (by Mitsubishi KagakuIatron, Inc.). Rats in the comparative groups were orally administeredmosapride (MOS) or metoclopropamide (MET), which are ordinarynausea/vomiting inhibitors. Each dose of MOS was 10 mg/kg, and that ofMET was 3 mg/kg. Twenty-five hours after the first administration,dosing of MOS and MET was repeated. The results are shown in FIG. 1. Therats in the control group were administered distilled water in place ofthe test substance.

The results show that Rikkunshi-to promoted the production of activeghrelin in fasted normal rats.

Test Example 2 Ghrelin production promotion test 2

The ghrelin production promoting effect was investigated in fasted ratsunder anticancer agent treatment. Seven-week-old SD male rats (8/group)were orally administered a test substance Rikkunshi-to (TJ-43, byTsumura & Co.) at a dose of 500 or 1000 mg/kg, and kept fasted for 24hours. Then, the anticancer agent cisplatin (CDDP) was intraperitoneallyadministered at a dose of 2 mg/kg. Nearly at the same time, Rikkunshi-toat the same previous dose was orally administered to the rats (2ndadministration). Rats of the comparative groups were administeredondansetron (ODS), which is an ordinary nausea/vomiting inhibitor, bysubcutaneous injection to the back of each rat, at a dose of 0.4mg/kg/dose; and 1.5 hours after the CDDP administration, the second dosewas administered. Besides, cisapride (CIS), SB242084 (SB) or MET wereused as comparative drugs. These were orally administered at a dose of 5mg/kg (CIS), 7 mg/kg (SB) or 10 mg/kg (MET), respectively, one hourafter the CDDP administration. In all the experiments, the rats weredecapitated two hours after the CDDP administration and blood wascollected to determine the concentration of ghrelin by ELISA. Theconcentration of active ghrelin was determined for each group, in thesame method as in Test Example 1. The results obtained with Rikkunshi-toand ODS are shown in FIG. 2; and those obtained with CIS, SB and MET areshown in FIG. 3. Rats in the normal control group were administeredphysiological saline instead of CDDP, and distilled water instead of thetest substance; those in the control group were administered CDDP, anddistilled water instead of the test substance.

In these results, active ghrelin significantly increased in bothRikkunshi-to (500 and 1000 mg/kg) groups. On the other hand, nosignificant difference was detected in the comparative groups.

Example 1 Production of Rikkunshi-to Extract

Cut crude drugs—4 kg ATRACTYLODIS LANCEAE RHIZOMA, 4 kg GINSENG RADIX, 4kg PINELLIAE TUBER, 4 kg POLIA, 2 kg ZIZYPHI FRUCTUS, 2 kg AURANTIINOBILIS PERICARPIUM, 1 kg GLYCYRRHIZAE RADIX and 0.5 kg ZINGIBERISRHIZOMA —were taken, blended, and added to about 12 volumes of purewater. Then, the mixture was heated up to 95 or 100° C. under stirring.Thereafter, this was extracted for 60 minutes and the extract wasprocessed for solid-liquid separation; then, the separated liquid wasconcentrated under reduced pressure. The concentrate was mixed withsucrose fatty acid ester added to a final concentration of 1.0% of thesolid content of the extract, and dried by spraying to obtain a driedpowder extract of Rikkunshi-to, 4.04 kg.

Example 2 Production of Granules

Lactose (3.4225 kg) and magnesium stearate (0.0375 kg) were added to andmixed with the dried powder extract of Rikkunshi-to (4.04 kg) obtainedin Example 1. The mixture was tabletted using a tabletting machineaccording to conventional methods, and then pulverized, granulated andsieved to obtain good granules.

INDUSTRIAL APPLICABILITY

The ghrelin production promoter of the invention can increase anacylated active ghrelin that has an activity of growth hormone secretioneffect and the like.

Accordingly, this is useful as a therapeutic agent for ghrelin-relateddwarfism; cardiac diseases such as heart failure and myocardialinfarction; and central nerve disorders such as Alzheimer's disease andParkinson's disease. In addition, the ghrelin production promoter of theinvention has an excellent safety profile, since the active ingredientis Rikkunshi-to which has been traditionally used as a prescription drugin Japan and China.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 Concentration of active ghrelin after administration of each testsubstance in Test Example 1.

FIG. 2 Concentration of active ghrelin in the Rikkunshi-to and ODSadministration groups in Test Example 2.

FIG. 3 Concentration of active ghrelin concentration in the CIS, SB andMET administration groups in Test Example 2.

1-6. (canceled)
 7. A method of promoting production of ghrelin in asubject in need thereof, the method comprising administering aneffective amount of Rikkunshi-to to the subject.
 8. A method ofpromoting secretion of a growth hormone in a subject in need thereof,the method comprising administering an effective amount of Rikkunshi-toto the subject to promote secretion of a growth hormone and promoteproduction of ghrelin.
 9. A method of treating a disease selected fromthe group consisting of myocardial infarction, heart failure,Alzheimer's disease and chronic obstructive pulmonary disease in asubject in need thereof, the method comprising administering aneffective amount of Rikkunshi-to to the subject to treat the disease andpromote production of ghrelin.
 10. The method of claim 9, wherein thedisease is myocardial infarction.
 11. The method of claim 9, wherein thedisease is heart failure.
 12. The method of claim 9, wherein the diseaseis chronic obstructive pulmonary disease.
 13. The method of claim 9,wherein the disease is Alzheimer's disease.